Awards and Professional Activities:
1999-2001: Hal and Alma Reagan Fellowship for academic excellence in the field of cancer research.
Distinguished Member of The National Society of Collegiate Scholars, 2006.Overview

Research in the Bothner lab is directed toward understanding biological function by investigating systems. This research takes us from the atomic scale provided by high resolution structural models of viruses to the complex interaction networks of nucleic acids, metabolites, and proteins that make up a living system. A diverse set of analytical, biophysical, biochemical, and cell biology techniques are used in the discovery process. Research interests include the assembly and stability of virus particles, extremophiles, metabolomics, proteomics, and transcriptomics. Specific projects under investigation are a system-wide analysis of the cell cellular response to stress of Sulfolobus solfataricus, metabolomic analysis of hemorrhagic shock, novel anti-Hepatitis B compounds, the use of Adeno Associated virus in gene therapy, systems biology of Ignicoccus-Nanoarchaeum mutualism.
The Bothner lab is part of the Center for Bio-Inspired Nanomaterials and the Thermal Biology Institute.

Keywords:
Proteomics, Chemical Biology, Biophysical, Biochemistry

Stress Response in an Extremophile

The idea that life is a delicate balance of chemical processes that can occur only within a narrow range of conditions is changing as scientists continue to discover life in extreme environments. The thermal features of Yellowstone National Park are one example. Pools of nearly boiling acid, once thought to be void of life, are now known to contain thriving populations of unicellular organisms and their viruses. Of the three domains of life (Eukarya, Bacteria, and Archaea), the Archaea are the least understood. Many of the organisms that are classified as extremophiles are members the archaeal domain of life. Currently these organisms are the focus of intense research because of our lack of understanding of their ability to thrive in conditions once thought uninhabitable and the possibility of isolating enzymes that can with stand harsh industrial conditions. The specific objectives of this project are two-fold: 1) learn about viruses from extreme environments. 2) understand the Sulfolbus solfataricus response to viral infection, oxidative stress, and heavy metals. Cutting edge proteomics, metabolomics, and activity-based protein profiling (ABPP) are being applied to these studies. Among the many exciting findings from this work is the extensive use of protein post-translational modification in Archaea. The relatively small genome size of Sulfolobus makes this an ideal organism for systems biology studies.
This is being pursued in conjunction with other MSU research groups within the Thermal Biology Institute.

It is now possible to investigate the entire population of a class of biomolecules in a cell or tissue simultaneously. Genomics, the study of all of the genes in an organism was the first such technology. Now, transcriptomics, proteomics, metabolomics, lipidomics, and a host of other â€œomicsâ€ techniques are changing the way biologists view and study cells. Data from these techniques are catalyzing fundamental changes in our understanding of biology. One such change is the view of organisms as dynamic systems composed of networks. These networks are highly interwoven and span molecular classes. Significantly, it is now clear that it is not possible to understand a biological system by only studying the parts. It is the goal of systems biology to integrate data on cellular networks into mathematical models with predictive power. While models that can predict biological responses have begun to emerge, they are still very limited. Our research focuses on model systems with the goal of developing biological models that can then be extended to more complex organisms and eventually human disease.
Archaea are philogenetic intermediates between Eukaryotes and Bacteria. The general lack of knowledge about their physiology and small genome size make them a good choice for systems biology studies. The extremophiles S. solfataricus is a member of the Archaeal domain of life and is a modle system for understanding adaptations to life in extreme environments. Using deep sequencing, proteomics, and metabolomics we are mapping metabolic pathways and building models of signaling networks in this interest organism that inhabits many of the hot springs in Yellow Stone National Park.
Few, if any, microbes live in functional or spatial isolation. The nature of the various types of inter-species interactions can be complex, ranging from competition to mutualism or syntrophy (metabolic interdependence). Such relationships can impact keystone species and play a major role in energy and element cycles at scales that extend past ecosystem boundaries. Currently, there is little understanding of the fundamental mechanisms of interspecies recognition and communication, how mutualism and syntrophy impacts microbial genome evolution and what genetic regulatory mechanisms control metabolic/energetic coupling between species in response to environmental factors. To address these questions we are studying the archaeal symbiotic system Ignicoccus hospitalis-Nanoarchaeum equitans to characterize mechanisms of interspecific recognition and communication, modulation of gene expression and metabolic processes triggered by the association. With a combined genomic complement of less than 2000 genes and an obligate chemolithoautotrophic metabolism that integrates adaptive traits and biochemical processes of interest to DOE, this system represents the simplest microbial association known in nature and allows fundamental system level investigations and modeling of symbiosis.
We have adopted this system and are using a two-level proteomics approach to elucidate pathways and proteins involved with MNV infection. At the systems level, 2D differential gel electrophoresis (2D-DIGE) is being applied to generate an overview of the global changes in the host proteome during infection. This is being complemented by activity-based screening of enzyme classes. Activity-based protein profiling (ABPP) is currently the only technique that can directly measure specific protein activity across a biological system.
Research Associate: Dr. Walid Maaty. Post Doctoral: Monika Tokmina-Lukaszewska Graduate students: Joshua Heinemann, Patricia Mathabe, and Michelle Tigges.

Keywords:
Proteomics, Chemical Biology, Biochemistry, Analytical

Protein Dynamics

The solution-phase protein motion that is part of a multi-component complex can not always be inferred from the three-dimensional structure. For example, in contrast to the still-life representation of viral capsids in models based on cryo-electron microscopy and X-ray crystallography, these supramolecular protein complexes are highly dynamic in solution. The range and frequency of capsid protein dynamics are poorly understood, despite evidence that the infectivity of animal viruses requires conformational freedom. Protein function is intimately connected to dynamics and therefore knowledge of the frequency, range, and coordination of motion by supramolecular complexes is critical to understanding how they function. Our lab uses viruses as a paradigm for studying protein dynamics in supramolecular complexes. A number of biophysical techniques including time-resolved fluorescence, differential scanning fluorimetry, hydrogen-deuterium exchange, kinetic hydrolysis, and quantitative mass spectrometry, we are determining the free energy and rates of large scale protein motion within viral particles. These are the first quantitative measurements for protein dynamics in megadalton complexes. Selective protein labeling, and quartz crystal microbalance measurements are a few of the additional methods applied to the quantitative analysis of virus particle stability and dynamics. Current projects include the use of Adeno Associated virus in gene therapy and characterization of a novel class of anti-Hepatitis B compounds.
Graduate students: Navid Movahed and Vamseedhar Rayaprolu.

Keywords:
Biochemistry, Biophysical, Chemical Biology, Structure

Protein Cages as Nanomaterials

Nature has evolved active bio-architectures that are both dynamic and responsive individually as well as collectively when assembled into hierarchical structures. In fact, dynamic protein regions are responsible for biological mineral nucleation, surface recognition, chemical reactivity, and targeting. The concerted protein motion that is part of a multi-component biomolecular complex is rarely obvious from the high resolution three-dimensional structure. Protein function is intimately connected to dynamics and therefore knowledge of the frequency, range, and coordination of motion by supramolecular complexes is critical to understanding function and the development of bio-inspired nanomaterials. The extremely large size and icosahedral architecture of virus capsids limit the use of many standard techniques for studying protein motion such as NMR and FRET. To overcome these problems, we employ an array of biophysical techniques to study the solution phase behavior of viruses. Kinetic hydrolysis, an approach being developed in our lab, is a straight-forward and powerful technique for identifying the dynamic regions within a single protein or in the context of a multi-component complex. Protein dynamics is being investigated at three levels: the dynamics of the subunit, the assembled cage architecture, and the dynamics associated with higher order particle/particle and surface/particle interactions. The long-term goal of this effort is to understand dynamics of the nanoparticle/cage system at each distinct level of complexity so that the underlying mechanism of nucleation, recognition, and functionality can be elucidated and exploited. This work is being conducted in collaboration with other research groups in the Center for Bio-Inspired Nanomaterials.
Graduate students: Navid Movahed and Vamseedhar Rayaprolu.

Hemorrhagic shock, a result of acute blood loss, is the third leading cause of death in the United States and the leading cause of death for people under 40. It is also a priming event for the development of Multiple Organ Dysfunction Syndrome (MODS). From a physiological stand point, decreased blood volume leads to a rapid decline in blood pressure, decreasing blood flow. An anaerobic environment in the peripheral tissues ensues that leads to a switch in cellular metabolism. For reasons that are not fully understood, dramatic physiologic and metabolic changes leading to clinical shock often occur. It is our hypothesis that metabolic changes arise early in the process of hemorrhage that predispose the patient to the course of events leading to the aforementioned complications. We are investigating changes at the level of the metabolome across time in a clinically relevant, large animal model of hemorrhagic shock with the goal of identification of biomarkers specific for risk of complications. To achieve this, state-of-the-art liquid chromatography mass spectrometry (LCMS) is employed for the purposes of identifying small molecules that change between treatment groups and across time. Speed, sensitivity, and reproducibility make mass spectrometry a powerful technique for use in metabolomics and we take advantage of the state of the art mass spectrometry, proteomics and metablomics facility in the Chemistry and Biochemistry Department in our research. Post Doctoral: Monika Tokmina-Lukaszewska.